CHARACTERIZATION OF HIGH-MOLECULAR-MASS LINEAR POLYACRYLAMIDE POWDER PREPARED BY EMULSION POLYMERIZATION AS A REPLACEABLE POLYMER MATRIX FOR DNA-SEQUENCING BY CAPILLARY ELECTROPHORESIS

In a previous paper, a 2% w/w replaceable high molecular mass linear p olyacrylamide solution (high molecular mass LPA) was used to achieve l ong read-lengths for DNA sequencing by capillary electrophoresis (E. C arrilho et al., Anal. Chem. 1996, 68, 3305-3313). In that work, the po lymer was prepared by polymerization in water at 6% w/w, followed by d ilution to 2% w/w. In this study, an improved method for preparation o f high molecular mass LPA was developed, based on inverse emulsion pol ymerization. With this polymerization procedure, the LPA results in a molecular mass of approximately 9 MDa, with characteristics of a fine powder of high purity and practically unlimited shelf life. Using size exclusion chromatography (SEC) and viscosity measurements to characte rize the polymer, good batch-to-batch reproducibility was found. It wa s observed that the viscous polymer solutions made from these high mol ecular mass polymers require careful preparation and handling because the method of dissolution could affect the molecular mass distribution and the resultant separation of DNA components. Solutions containing 2% w/w of LPA made by emulsion polymerization were simple to prepare, resulting in excellent performance as a replaceable matrix for DNA seq uencing by capillary electrophoresis. The viscosity of the polymer dec reased exponentially when pressure was applied, allowing easy replacem ent from a capillary using a syringe. With a properly prepared matrix, a read-length of more than 1000 bases in 80 min with an accuracy bett er than 97%, and better than 99% for the first 800 bases, could be ach ieved.